1. Field of the Invention
The present invention relates to an aldolase and a process for producing substituted α-keto acid, and more particularly, to an aldolase that may be preferably used in the synthesis of 4-(indol-3-ylmethyl)-4-hydroxy-2-oxoglutarate (hereinafter, “IHOG”), which is useful as an intermediate in monatin synthesis, and a process for producing substituted α-keto acids.
2. Discussion of the Background
Monatin, which has the structure shown in formula (5) below, a naturally sweet amino acid that is isolated and extracted from the roots of shrubs in Southern Africa. It has a potent sweet taste equivalent to several ten to several thousand times that of sucrose, and is expected to be useful as a sweetener. However, the usefulness of monatin has only been recently discovered, and a process for synthesizing monatin as the level of industrial production has yet to be established.

Under these circumstances, the inventors of the present invention developed a novel process for producing monatin composed of the following reactions (1) and (2) by using indole pyruvic acid and pyruvic acid which may be acquired as reagents.
(1) A reaction step of synthesizing precursor keto acid (IHOG) by aldol condensation of indole pyruvic acid and pyruvic acid (and/or oxaloacetic acid); and,
(2) a reaction step of aminating the second position of IHOG.

There have been no previous reports of examples of synthesizing precursor keto acid (IHOG) from indole pyruvic acid and pyruvic acid (or oxaloacetic acid) using a microbial enzyme system for the aldol condensation reaction of (1) in the aforementioned synthesis route of monatin.
Examples of microbial enzymes that catalyze aldol condensation using two molecules of α-keto acid (or substituted α-keto acid) as a substrate reported thus far include 4-hydroxy-4-methyl-2-oxoglutarate aldolase derived from bacteria belonging to the genus Pseudomonas, and 4-hydroxy-2-oxoglutarate aldolase present in E. coli, B. subtilis, and so forth.
The former 4-hydroxy-4-methyl-2-oxoglutarate aldolase has been reported to catalyze a reaction in which 4-hydroxy-4-methyl-2-oxoglutarate (4-HMG) is formed from two molecules of pyruvic acid, and a reaction in which one molecule of oxaloacetic acid and one molecule of pyruvic acid are formed from 4-oxalocitramalate (Kiyofumi Maruyama, Journal of Biochemistry, 108, 327-333 (1990)). Furthermore, the latter 4-hydroxy-2-oxoglutarate aldolase is known to catalyze a reaction in which 4-hydroxy-2-oxoglutarate (4HG) is formed from one molecule of glyoxylic acid and one molecule of pyruvic acid.
However, there have been no reports or findings indicating that any of these microorganisms are associated with activity that cleaves 4-phenylmethyl-4-hydroxy-2-oxoglutarate (PHOG) or activity that synthesizes a precursor keto acid (IHOG) of monatin from indole pyruvic acid and pyruvic acid (or oxaloacetic acid), and whether or not the aldolases produced by these microbial strains may be used in the aforementioned synthesis route of monatin is unknown.